Combination of (a) a phosphoinositide 3-kinase inhibitor and (b) a modulator of ras/raf/mek pathway

ABSTRACT

The invention relates to a pharmaceutical combination which comprises (a) a phosphoinositide 3-kinase inhibitor compound and (b) a compound which modulates the Ras/Raf/Mek pathway for the treatment of a proliferative disease, especially a solid tumor disease; a pharmaceutical composition comprising such a combination; the use of such a combination for the preparation of a medicament for the treatment of a proliferative disease; a commercial package or product comprising such a combination as a combined preparation for simultaneous, separate or sequential use; and to a method of treatment of a warm-blooded animal, especially a human.

The invention relates to a pharmaceutical combination which comprises(a) a phosphoinositide 3-kinase (PI3K) inhibitor compound and (b) acompound which modulates the Ras/Raf/Mek pathway and optionally at leastone pharmaceutically acceptable carrier for simultaneous, separate orsequential use, in particular for the treatment of a proliferativedisease, especially a proliferative disease in which the Ras/Raf/MeK andPI3K/Akt pathways are concomitantly dysregulated; a pharmaceuticalcomposition comprising such a combination; the use of such a combinationfor the preparation of a medicament for the treatment of a proliferativedisease; a commercial package or product comprising such a combinationas a combined preparation for simultaneous, separate or sequential use;and to a method of treatment of a warm-blooded animal, especially ahuman.

WO2006/122806 describes imidazoquinoline derivatives, which have beendescribed to inhibit the activity of lipid kinases, such as PI3-kinases.Specific imidazoquinoline derivatives which are suitable for the presentinvention, their preparation and suitable pharmaceutical formulationscontaining the same are described in WO2006/122806 and include compoundsof formula I

wherein

R₁ is naphthyl or phenyl wherein said phenyl is substituted by one ortwo substituents independently selected from the group consisting ofHalogen; lower alkyl unsubstituted or substituted by halogen, cyano,imidazolyl or triazolyl; cycloalkyl; amino substituted by one or twosubstituents independently selected from the group consisting of loweralkyl, lower alkyl sulfonyl, lower alkoxy and lower alkoxy loweralkylamino; piperazinyl unsubstituted or substituted by one or twosubstituents independently selected from the group consisting of loweralkyl and lower alkyl sulfonyl; 2-oxo-pyrrolidinyl; lower alkoxy loweralkyl; imidazolyl; pyrazolyl; and triazolyl;

R₂ is O or S;

R₃ is lower alkyl;

R₄ is pyridyl unsubstituted or substituted by halogen, cyano, loweralkyl, lower alkoxy or piperazinyl unsubstituted or substituted by loweralkyl; pyrimidinyl unsubstituted or substituted by lower alkoxy;quinolinyl unsubstituted or substituted by halogen; quinoxalinyl; orphenyl substituted with alkoxy

R₅ is hydrogen or halogen;

n is 0 or 1;

R₆ is oxido;

with the proviso that if n=1, the N-atom bearing the radical R₆ has apositive charge;

R₇ is hydrogen or amino;

or a tautomer thereof, or a pharmaceutically acceptable salt, or ahydrate or solvate thereof.

The radicals and symbols as used in the definition of a compound offormula I have the meanings as disclosed in WO2006/122806 whichpublication is hereby incorporated into the present application byreference.

A preferred compound of the present invention is a compound which isspecifically described in WO2006/122806. A very preferred compound ofthe present invention is2-methyl-2-[4-(3-methyl-2-oxo-8-quinolin-3-yl-2,3-dihydro-imidazo[4,5-c]quinolin-1-yl)-phenyl]-propionitrileand its monotosylate salt (COMPOUND A). The synthesis of2-methyl-2-[4-(3-methyl-2-oxo-8-quinolin-3-yl-2,3-dihydro-imidazo[4,5-c]quinolin-1-yl)-phenyl]-propionitrileis for instance described in WO2006/122806 as Example 1. Another verypreferred compound of the present invention is8-(6-methoxy-pyridin-3-yl)-3-methyl-1-(4-piperazin-1-yl-3-trifluoromethyl-phenyl)-1,3-dihydro-imidazo[4,5-c]quinolin-2-one(COMPOUND B). The synthesis of8-(6-methoxy-pyridin-3-yl)-3-methyl-1-(4-piperazin-1-yl-3-trifluoromethyl-phenyl)-1,3-dihydro-imidazo[4,5-c]quinolin-2-oneis for instance described in WO2006/122806 as Example 86.

WO07/084,786 describes pyrimidine derivatives, which have been found theactivity of lipid kinases, such as PI3-kinases. Specific pyrimidinederivatives which are suitable for the present invention, theirpreparation and suitable pharmaceutical formulations containing the sameare described in WO07/084,786 and include compounds of formula I

-   -   or a stereoisomer, tautomer, or pharmaceutically acceptable salt        thereof, wherein, W is CR_(w) or N, wherein R_(w) is selected        from the group consisting of    -   (1) hydrogen,    -   (2) cyano,    -   (3) halogen,    -   (4) methyl,    -   (5) trifluoromethyl,    -   (6) sulfonamido;    -   R₁ is selected from the group consisting of    -   (1) hydrogen,    -   (2) cyano,    -   (3) nitro,    -   (4) halogen,    -   (5) substituted and unsubstituted alkyl,    -   (6) substituted and unsubstituted alkenyl,    -   (7) substituted and unsubstituted alkynyl,    -   (8) substituted and unsubstituted aryl,    -   (9) substituted and unsubstituted heteroaryl,    -   (10) substituted and unsubstituted heterocyclyl,    -   (11) substituted and unsubstituted cycloalkyl,    -   (12) —COR_(1a),    -   (13) —CO₂R_(1a),    -   (14) —CONR_(1a)R_(1b),    -   (15) —NR_(1a)R_(1b),    -   (16) —NR_(1a)COR_(1b),    -   (17) —NR_(1a)SO₂R_(1b),    -   (18) —OCOR_(1a),    -   (19) —OR_(1a),    -   (20) —SR_(1a),    -   (21) —SOR_(1a),    -   (22) —SO₂R_(1a), and    -   (23) —SO₂NR_(1a)R_(1b),    -   wherein R_(1a), and R_(1b) are independently selected from the        group consisting of    -   (a) hydrogen,    -   (b) substituted or unsubstituted alkyl,    -   (c) substituted and unsubstituted aryl,    -   (d) substituted and unsubstituted heteroaryl,    -   (e) substituted and unsubstituted heterocyclyl, and    -   (f) substituted and unsubstituted cycloalkyl;    -   R₂ is selected from the group consisting    -   (1) hydrogen,    -   (2) cyano,    -   (3) nitro,    -   (4) halogen,    -   (5) hydroxy,    -   (6) amino,    -   (7) substituted and unsubstituted alkyl,    -   (8) —COR_(2a), and    -   (9) —NR_(2a)COR_(2b),    -   wherein R_(2a), and R_(2b) are independently selected from the        group consisting of    -   (a) hydrogen, and    -   (b) substituted or unsubstituted alkyl;    -   R₃ is selected from the group consisting of    -   (1) hydrogen,    -   (2) cyano,    -   (3) nitro,    -   (4) halogen,    -   (5) substituted and unsubstituted alkyl,    -   (6) substituted and unsubstituted alkenyl,    -   (7) substituted and unsubstituted alkynyl,    -   (8) substituted and unsubstituted aryl,    -   (9) substituted and unsubstituted heteroaryl,    -   (10) substituted and unsubstituted heterocyclyl,    -   (11) substituted and unsubstituted cycloalkyl,    -   (12) —COR_(3a),    -   (13) —NR_(3a)R_(3b),    -   (14) —NR_(3a)COR_(3b),    -   (15) —NR_(3a)SO₂R_(3b),    -   (16) —OR_(3a),    -   (17) —SR_(3a),    -   (18) —SOR_(3a),    -   (19) —SO₂R_(3a), and    -   (20) —SO₂NR_(3a)R_(3b),    -   wherein R_(3a), and R_(3b) are independently selected from the        group consisting of    -   (a) hydrogen,    -   (b) substituted or unsubstituted alkyl,    -   (c) substituted and unsubstituted aryl,    -   (d) substituted and unsubstituted heteroaryl,    -   (e) substituted and unsubstituted heterocyclyl, and    -   (f) substituted and unsubstituted cycloalkyl; and    -   R₄ is selected from the group consisting of    -   (1) hydrogen, and    -   (2) halogen.

The radicals and symbols as used in the definition of a compound offormula I have the meanings as disclosed in WO07/084,786 whichpublication is hereby incorporated into the present application byreference.

A preferred compound of the present invention is a compound which isspecifically described in WO07/084,786. A very preferred compound of thepresent invention is5-(2,6-di-morpholin-4-yl-pyrimidin-4-yl)-4-trifluoromethyl-pyridin-2-ylamine(COMPOUND C). The synthesis of5-(2,6-di-morpholin-4-yl-pyrimidin-4-yl)-4-trifluoromethyl-pyridin-2-ylamineis described in WO07/084,786 as Example 10.

The Ras/Raf/Mek signal transduction pathway is activated through theinitial extracellular binding and stimulation of RTKs by theirrespective cognate ligands. Upon autophosphorylation of specifc tyrosineresidues in the cytosolic domain of RTKs, the Grb2-Sos complextranslocates to the plasma membrane, and converts the inactive Ras·GDPto active Ras·GTP. The interaction between Grb2 and the activatedkinases or the phosphorylated receptor associated proteins is mediatedby the SH2 domain of the signalling protein that recognizes specificphosphotyrosine sequences. Ras undergoes a conformational changes uponGTP binding and causes the recruitment of Raf-1 to the cytoplasmicmembrane where is phosphorylated by several kinases and simultaneousdisphosphorylated at key residues by protein phosphatase-2B. ActivatedRaf phosphorylates Mek on two serine residues in the activation loop,which results in the activation of this protein kinase. Mek thenphosphorylates and activates Erk, allowing its translocation to thenucleus where it phosphorylates transcriptional factors permitting theexpression of a variety of genes. As shown in this section, severalcomponents of the Raf/Mek/Erk cascade represent unique opportunities forcancer treatment, e.g. genetic alterations such as HRAS, KRAS, NRAS orBRAF mutations or gene amplifications.

Sorafenib, also known as Nexavar™ or BAY 43-9006, has been the first Rafkinase modulator to receive marketing approval for treatment of advancedrenal cell carcinoma (RCC). Sorafenib is a bi-aryl urea derivative ofthe following structure:

Sorafenib and methods for its preparation have been described e.g. inU.S. Pat. No. 7,235,576 or U.S. Pat. No. 7,351,834.

RAF265 (previously known as CHIR-265) is another orally bioavailable Rafmodulator of following structure:

RAF265 and methods for its preparation for are for instance described inWO07/030,377.

Further Raf kinase modulators include for instance SB590885, XL281,PLX4032.

Mek kinase modulators include compounds described in WO02/06213, inparticular compound PD-0325901 which has following structure:

Another related Mek kinase inhibitor is compound PD-181461.

Further Mek kinase modulators include compounds described inWO03/077914, in particular compound ARRY1428861/AZD6244 of structure

and ARRY-509 and the compounds described in WO05/051906, WO05/023251,WO03/077855 and US20050049419 covering N3-alkylated benzimidazoles andother similar heterocyclic derivatives as MEK inhibitors for thetreatment of hyperproliferative diseases.

Further Mek kinase modulators include for instance XL518, JTP-74057,AS-701255, AS-701173.

Hence, the present invention also pertains to a combination such as acombined preparation or a pharmaceutical composition which comprises (a)a phosphoinositide 3-kinase (PI3K) inhibitor compound and (b) a compoundwhich modulates the Ras/Raf/Mek pathway. More particularly, in a firstembodiment, the present invention relates to a combination whichcomprises (a) a phosphoinositide 3-kinase (PI3K) inhibitor compound and(b) a Raf kinase modulator or Mek kinase modulator.

The term “a combined preparation”, as used herein defines especially a“kit of parts” in the sense that the combination partners (a) and (b) asdefined above can be dosed independently or by use of different fixedcombinations with distinguished amounts of the combination partners (a)and (b), i.e. simultaneously or at different time points. The parts ofthe kit of parts can then, e.g., be administered simultaneously orchronologically staggered, that is at different time points and withequal or different time intervals for any part of the kit of parts. Theratio of the total amounts of the combination partner (a) to thecombination partner (b) to be administered in the combined preparationcan be varied, e.g. in order to cope with the needs of a patientsub-population to be treated or the needs of the single.

It has been found that combination therapy with an PI3K inhibitor and araf-kinase or mek-kinase inhibitor results in unexpected improvement inthe treatment of tumor diseases. When administered simultaneously,sequentially or separately, the PI3K inhibitor and the raf-kinase ormek-kinase inhibitor interact in a synergistic manner to inhibit cellproliferation. This unexpected synergy allows a reduction in the doserequired of each compound, leading to a reduction in the side effectsand enhancement of the clinical effectiveness of the compounds andtreatment.

Determining a synergistic interaction between one or more components,the optimum range for the effect and absolute dose ranges of eachcomponent for the effect may be definitively measured by administrationof the components over different w/w ratio ranges and doses to patientsin need of treatment. For humans, the complexity and cost of carryingout clinical studies on patients renders impractical the use of thisform of testing as a primary model for synergy. However, the observationof synergy in one species can be predictive of the effect in otherspecies and animal models exist, as described herein, to measure asynergistic effect and the results of such studies can also be used topredict effective dose and plasma concentration ratio ranges and theabsolute doses and plasma concentrations required in other species bythe application of pharmacokinetic/pharmacodynamic methods. Establishedcorrelations between tumor models and effects seen in man suggest thatsynergy in animals may e.g. be demonstrated in the BN472 tumor model orpancreatic PANC1 and MiaPaCa2 tumor models as described in the Examplesbelow.

In one aspect the present invention provides a synergistic combinationfor human administration comprising (a) PI3K inhibitor compound and (b)a compound which modulates the Ras/Raf/Mek pathway, or pharmaceuticallyacceptable salts or solvates thereof, in a combination range (w/w) whichcorresponds to the ranges observed in a tumor model, e.g. as describedin the Examples below, used to identify a synergistic interaction.Suitably, the ratio range in humans corresponds to a non-human rangeselected from between 50:1 to 1:50 parts by weight, 50:1 to 1:20, 50:1to 1:10, 50:1 to 1:1, 20:1 to 1:50, 20:1 to 1:20, 20:1 to 1:10, 20:1 to1:1, 10:1 to 1:50, 10:1 to 1:20, 10:1 to 1:10, 10:1 to 1:1, 1:1 to 1:50,1.1 to 1:20 and 1:1 to 1:10. More suitably, the human range correspondsto a non-human range of the order of 10:1 to 1:1 or 5:1 to 1:1 or 2:1 to1:1 parts by weight.

According to a further aspect, the present invention provides asynergistic combination for administration to humans comprising an (a) aPI3K inhibitor compound and (b) a compound which modulates theRas/Raf/Mek pathway or pharmaceutically acceptable salts thereof, wherethe dose range of each component corresponds to the synergistic rangesobserved in a suitable tumor model, e.g. the tumor models described inthe Examples below, primarily used to identify a synergisticinteraction. Suitably, the dose range of the PI3K inhibitor compound inhuman corresponds to a dose range of 1-50 mg/kg, more suitably 1-30mg/kg (e.g. 1-35 mg/kg or 1-10 mg/kg for Compound A, 1-25 mg/kg forCompound B) in a suitable tumor model, e.g. a rat or mouse model asdescribed in the Examples below.

For the compound which modulates the Ras/Raf/Mek pathway, the dose rangein the human suitably corresponds to a synergistic range of 1-50 mg/kgor 1-30 mg/kg (e.g. 1-25 mg/kg, 1-10 mg/kg or 1-2.5 mg/kg) in a suitabletumor model, e.g. a rat or mouse model as described in the Examplesbelow.

Suitably, the dose of PI3K inhibitor compound for use in a human is in arange selected from 1-1200 mg, 1-500 mg, 1-100 mg, 1-50 mg, 1-25 mg,500-1200 mg, 100-1200 mg, 100-500 mg, 50-1200 mg, 50-500 mg, or 50-100mg, suitably 50-100 mg, once daily or twice daily (b.i.d.) or threetimes per day (t.i.d.), and the dose of compound which modulates theRas/Raf/Mek pathway is in a range selected from 1-1000 mg, 1-500 mg,1-200 mg, 1-100 mg, 1-50 mg, 1-25 mg, 10-100 mg, 10-200 mg, 50-200 mg or100-500 mg once daily, b.i.d or t.i.d.

In accordance with a further aspect the present invention provides asynergistic combination for administration to humans comprising an (a) aPI3K inhibitor compound at 10%-100%, preferably 50%-100% or morepreferably 70%-100%, 80%-100% or 90%-100% of the maximal tolerable dose(MTD) and (b) a compound which modulates the Ras/Raf/Mek pathway at10%-100%, preferably 50%-100% or more preferably 70%-100%, 80%-100% or90%-100% of the MTD. In a preferred embodiment one of the compounds,preferably the PI3K inhibitor compound, is dosed at the MTD and theother compound, preferably the compound which modulates the Ras/Raf/Mekpathway, is dosed at 50%-100% of the MTD, preferably at 60%-90% of theMTD. The MTD corresponds to the highest dose of a medicine that can begiven without unacceptable side effects. It is within the art todetermine the MTD. For instance the MTD can suitably be determined in aPhase I study including a dose escalation to characterize dose limitingtoxicities and determination of biologically active tolerated doselevel.

In one embodiment of the invention, (a) the phosphoinositide 3-kinase(PI3K) inhibitor compound inhibitor is selected from the groupconsisting of COMPOUND A, COMPOUND B or COMPOUND C.

In one embodiment of the invention, (b) the Raf kinase modulator isselected from the group consisting of Sorafenib, Raf265, SB590885, XL281and PLX4032. In another embodiment, (b) the Mek kinase modulator isselected from the group consisting of PD325901, PD-181461,ARRY142886/AZD6244, ARRY-509, XL518, JTP-74057, AS-701255, AS-701173,AZD8330, ARRY162, ARRY300, RDEA436, E6201, RO4987655/R-7167, GSK1120212and AS703026.

The term “treating” or “treatment” as used herein comprises the atreatment effecting a delay of progression of a disease. The term “delayof progression” as used herein means administration of the combinationto patients being in a pre-stage or in an early phase of theproliferative disease to be treated, in which patients for example apre-form of the corresponding disease is diagnosed or which patients arein a condition, e.g. during a medical treatment or a condition resultingfrom an accident, under which it is likely that a corresponding diseasewill develop.

In one embodiment of the present invention, the proliferative disease ismelanoma, lung cancer, colorectal cancer (CRC), breast cancer, kidneycancer such as e.g. renal cell carcinoma (RCC), liver cancer, acutemyelogenous leukemia (AML), myelodysplastic Syndromes (MDS),non-small-cell lung cancer (NSCLC), thyroid cancer, pancreatic cancer,neurofibromatosis or hepatocellular carcinoma.

In another embodiment of the present invention, the proliferativedisease is a solid tumor. The term “solid tumor” especially means breastcancer, ovarian cancer, cancer of the colon and generally the GI(gastro-intestinal) tract, cervix cancer, lung cancer, in particularsmall-cell lung cancer, and non-small-cell lung cancer, head and neckcancer, bladder cancer, cancer of the prostate or Kaposi's sarcoma. Thepresent combination inhibits the growth of solid tumors, but also liquidtumors. Furthermore, depending on the tumor type and the particularcombination used a decrease of the tumor volume can be obtained. Thecombinations disclosed herein are also suited to prevent the metastaticspread of tumors and the growth or development of micrometastases. Thecombinations disclosed herein are in particular suitable for thetreatment of poor prognosis patients, especially such poor prognosispatients having metastatic melanome or pancreatic cancer.

The cancer to be treated can have a genetic alteration in theRas/Raf/Mek signal transduction pathway such as e.g. a HRAS, KRAS, NRASor BRAF mutation or gene amplification. In one embodiment the cancer tobe treated has a KRAS mutation, e.g. KRAS mutated pancreas cancer, coloncancer, lung cancer (e.g. NSCLC) or leukemias.

The structure of the active agents identified by code nos., generic ortrade names may be taken from the actual edition of the standardcompendium “The Merck Index” or from databases, e.g. PatentsInternational (e.g. IMS World Publications). The corresponding contentthereof is hereby incorporated by reference.

It will be understood that references to the combination partners (a)and (b) are meant to also include the pharmaceutically acceptable salts.If these combination partners (a) and (b) have, for example, at leastone basic center, they can form acid addition salts. Corresponding acidaddition salts can also be formed having, if desired, an additionallypresent basic center. The combination partners (a) and (b) having anacid group (for example COOH) can also form salts with bases. Thecombination partner (a) or (b) or a pharmaceutically acceptable saltthereof may also be used in form of a hydrate or include other solventsused for crystallization.

A combination which comprises (a) a phosphoinositide 3-kinase inhibitorcompound and (b) a compound which modulates the Ras/Raf/Mek pathway, inwhich the active ingredients are present in each case in free form or inthe form of a pharmaceutically acceptable salt and optionally at leastone pharmaceutically acceptable carrier, will be referred to hereinafteras a COMBINATION OF THE INVENTION.

The COMBINATION OF THE INVENTION are both synergistic and additiveadvantages, both for efficacy and safety. Therapeutic effects ofcombinations of a phosphoinositide 3-kinase inhibitor compound with acompound which modulates the Ras/Raf/Mek pathway can result in lowersafe dosages ranges of each component in the combination.

The pharmacological activity of a COMBINATION OF THE INVENTION may, forexample, be demonstrated in a clinical study or in a test procedure asessentially described hereinafter. Suitable clinical studies are, forexample, open label non-randomized, dose escalation studies in patientswith advanced solid tumors. Such studies can prove the additive orsynergism of the active ingredients of the COMBINATIONS OF THEINVENTION. The beneficial effects on proliferative diseases can bedetermined directly through the results of these studies or by changesin the study design which are known as such to a person skilled in theart. Such studies are, in particular, suitable to compare the effects ofa monotherapy using the active ingredients and a COMBINATION OF THEINVENTION. Preferably, the combination partner (a) is administered witha fixed dose and the dose of the combination partner (b) is escalateduntil the Maximum Tolerated Dosage is reached.

It is one objective of this invention to provide a pharmaceuticalcomposition comprising a quantity, which is therapeutically effectiveagainst a proliferative disease comprising the COMBINATION OF THEINVENTION. In this composition, the combination partners (a) and (b) canbe administered together, one after the other or separately in onecombined unit dosage form or in two separate unit dosage forms. The unitdosage form may also be a fixed combination.

The pharmaceutical compositions according to the invention can beprepared in a manner known per se and are those suitable for enteral,such as oral or rectal, and parenteral administration to mammals(warm-blooded animals), including man. Alternatively, when the agentsare administered separately, one can be an enteral formulation and theother can be administered parenterally.

The novel pharmaceutical composition contain, for example, from about10% to about 100%, preferably from about 20% to about 60%, of the activeingredients. Pharmaceutical preparations for the combination therapy forenteral or parenteral administration are, for example, those in unitdosage forms, such as sugar-coated tablets, tablets, capsules orsuppositories, and furthermore ampoules. If not indicated otherwise,these are prepared in a manner known per se, for example by means ofconventional mixing, granulating, sugar-coating, dissolving orlyophilizing processes. It will be appreciated that the unit content ofa combination partner contained in an individual dose of each dosageform need not in itself constitute an effective amount since thenecessary effective amount can be reached by administration of aplurality of dosage units.

In preparing the compositions for oral dosage form, any of the usualpharmaceutical media may be employed, such as, for example, water,glycols, oils, alcohols, flavoring agents, preservatives, coloringagents; or carriers such as starches, sugars, microcrystallinecellulose, diluents, granulating agents, lubricants, binders,disintegrating agents and the like in the case of oral solidpreparations such as, for example, powders, capsules and tablets, withthe solid oral preparations being preferred over the liquidpreparations. Because of their ease of administration, tablets andcapsules represent the most advantageous oral dosage unit form in whichcase solid pharmaceutical carriers are obviously employed.

In particular, a therapeutically effective amount of each of thecombination partner of the COMBINATION OF THE INVENTION may beadministered simultaneously or sequentially and in any order, and thecomponents may be administered separately or as a fixed combination. Forexample, the method of delay of progression or treatment of aproliferative disease according to the invention may comprise (i)administration of the first combination partner in free orpharmaceutically acceptable salt form and (ii) administration of thesecond combination partner in free or pharmaceutically acceptable saltform, simultaneously or sequentially in any order, in jointlytherapeutically effective amounts, preferably in synergisticallyeffective amounts. The individual combination partners of theCOMBINATION OF THE INVENTION can be administered separately at differenttimes during the course of therapy or concurrently in divided or singlecombination forms. Furthermore, the term administering also encompassesthe use of a pro-drug of a combination partner that convert in vivo tothe combination partner as such. The instant invention is therefore tobe understood as embracing all such regimes of simultaneous oralternating treatment and the term “administering” is to be interpretedaccordingly.

The COMBINATION OF THE INVENTION can be a combined preparation or apharmaceutical composition.

Moreover, the present invention relates to a method of treating awarm-blooded animal having a proliferative disease comprisingadministering to the animal a COMBINATION OF THE INVENTION in a quantitywhich is therapeutically effective against said proliferative disease.

Furthermore, the present invention pertains to the use of a COMBINATIONOF THE INVENTION for the treatment of a proliferative disease and forthe preparation of a medicament for the treatment of a proliferativedisease.

Moreover, the present invention provides a commercial package comprisingas active ingredients COMBINATION OF THE INVENTION, together withinstructions for simultaneous, separate or sequential use thereof in thedelay of progression or treatment of a proliferative disease.

Preferred embodiments of the invention are represented by combinationscomprising

-   -   COMPOUND A, COMPOUND B or COMPOUND C and Sorafenib,    -   COMPOUND A, COMPOUND B or COMPOUND C and RAF265

In another preferred embodiment, the invention provides combinationscomprising

-   -   COMPOUND A and Sorafenib, Raf265, SB590885, XL281 or PLX4032.    -   COMPOUND B and Sorafenib, Raf265, SB590885, XL281 or PLX4032.    -   COMPOUND C and Sorafenib, Raf265, SB590885, XL281 or PLX4032.    -   COMPOUND A and PD325901, PD-181461, ARRY142886/AZD6244,        ARRY-509, XL518, JTP-74057, AS-701255, AS-701173, AZD8330,        ARRY162, ARRY300, RDEA436, E6201, RO4987655/R-7167, GSK1120212        or AS703026.    -   COMPOUND B and PD325901, PD-181461, ARRY142886/AZD6244,        ARRY-509, XL518, JTP-74057, AS-701255, AS-701173, AZD8330,        ARRY162, ARRY300, RDEA436, E6201, RO4987655/R-7167, GSK1120212        or AS703026.    -   COMPOUND C and PD325901, PD-181461, ARRY142886/AZD6244,        ARRY-509, XL518, JTP-74057, AS-701255, AS-701173, AZD8330,        ARRY162, ARRY300, RDEA436, E6201, RO4987655/R-7167, GSK1120212        or AS703026.

In further aspects, the present inventions provides

-   -   a combination which comprises (a) a COMBINATION OF THE        INVENTION, wherein the active ingredients are present in each        case in free form or in the form of a pharmaceutically        acceptable salt or any hydrate thereof, and optionally at least        one pharmaceutically acceptable carrier; for simultaneous,        separate or sequential use;    -   a pharmaceutical composition comprising a quantity which is        jointly therapeutically effective against a proliferative        disease of a COMBINATION OF THE INVENTION and at least one        pharmaceutically acceptable carrier;    -   the use of a COMBINATION OF THE INVENTION for the treatment of a        proliferative disease;    -   the use of a COMBINATION OF THE INVENTION for the preparation of        a medicament for the treatment of a proliferative disease;    -   the use of a combination COMBINATION OF THE INVENTION wherein        the PI3K inhibitor is selected from COMPOUND A, COMPOUND B or        COMPOUND C;    -   the use of a COMBINATION OF THE INVENTION wherein the compound        which modulates the Ras/Raf/Mek pathway is compound which        modulates Raf kinase activity, e.g. Sorafenib, RAF265, SB590885,        XL281, PLX4032; and    -   the use of COMBINATION OF THE INVENTION wherein the compound        which modulates the Ras/Raf/Mek pathway is a compound which        modulates Mek kinase activity, e.g. PD-0325901, PD-181461,        ARRY142886/AZD6244, ARRY-509, XL518, JTP-74057, AS-701255,        AS-701173, AZD8330, ARRY162, ARRY300, RDEA436, E6201,        RO4987655/R-7167, GSK1120212 or AS703026.

Moreover, in particular, the present invention relates to a combinedpreparation, which comprises (a) one or more unit dosage forms of aphosphoinositide 3-kinase inhibitor compound and (b) a compound whichmodulates the Ras/Raf/Mek pathway.

Furthermore, in particular, the present invention pertains to the use ofa combination comprising (a) a phosphoinositide 3-kinase inhibitorcompound and (b) a compound which modulates the Ras/Raf/Mek pathway forthe preparation of a medicament for the treatment of a proliferativedisease.

The effective dosage of each of the combination partners employed in theCOMBINATION OF THE INVENTION may vary depending on the particularcompound or pharmaceutical composition employed, the mode ofadministration, the condition being treated, the severity of thecondition being treated. Thus, the dosage regimen the COMBINATION OF THEINVENTION is selected in accordance with a variety of factors includingthe route of administration and the renal and hepatic function of thepatient. A physician, clinician or veterinarian of ordinary skill canreadily determine and prescribe the effective amount of the singleactive ingredients required to prevent, counter or arrest the progressof the condition. Optimal precision in achieving concentration of theactive ingredients within the range that yields efficacy withouttoxicity requires a regimen based on the kinetics of the activeingredients' availability to target sites.

When the combination partners employed in the COMBINATION OF THEINVENTION are applied in the form as marketed as single drugs, theirdosage and mode of administration can take place in accordance with theinformation provided on the package insert of the respective marketeddrug in order to result in the beneficial effect described herein, ifnot mentioned herein otherwise.

COMPOUND A may be administered to a human in a dosage range varying fromabout 50 to 800 mg/day.

COMPOUND B may be administered to a human in a dosage range varying fromabout 25 to 800 mg/day.

COMPOUND C may be administered to a human in a dosage range varying fromabout 25 to 800 mg/day.

Sorafenib may be administered to a human in a dosage range varying fromabout 75 to 800 mg/day, more preferable 400 mg taken twice daily.

The following Examples illustrate the invention described above; theyare not, however, intended to limit the scope of the invention in anyway. The beneficial effects of the COMBINATION OF THE INVENTION can alsobe determined by other test models known as such to the person skilledin the pertinent art.

FIGURE LEGEND

FIG. 1: A549 cells were incubated either with compound A, PD03251901(250 nM) alone or in combination for 30 min. Cells were then harvestedand estimation of phosphorylated levels of either Akt; p70S6K or ERKdone by regular Western-blotting.

FIG. 2: Rat bearing BN472 tumors were treated with the indicatedcompound at the indicated dose and schedule, either alone or incombination. Evolution of tumor growth were followed by calliperingthroughout the course of the study. *, P<0.05 (Dunnett's).

FIG. 3: Rat bearing BN472 tumors were treated with the indicatedcompound at the indicated dose and schedule, either alone or incombination. Evolution of body weight were followed throughout thecourse of the study. *, P<0.05 (Dunnett's).

FIG. 4: KRAS mutant PANC1 cells were grown subcutaneously on the flankof Harlan nude mice and treated with either compound A, AZD6244 alone orin combination for 12 days. Evolutions of Tumor volume (A) and bodyweight (B) were measured throughout the course of the study. *, p<0.05(Dunnett's).

FIG. 5: KRAS mutant MiaPaCa2 cells were grown subcutaneously on theflank of Harlan nude mice and treated with either compound A, AZD6244alone or in combination for 12 days. Evolutions of Tumor volume (A) andbody weight (B) were measured throughout the course of the study. *,p<0.05 (Dunnett's).

FIG. 6: KRAS mutant MiaPaCa2 cells were grown subcutaneously on theflank of Harlan nude mice and treated with either5-(2,6-di-morpholin-4-yl-pyrimidin-4-yl)-4-trifluoromethyl-pyridin-2-ylamine(COMPOUND C, termed here cpd B), AZD6244 alone or in combination for 12days. Evolutions of Tumor volume (A) and body weight (B) were measuredthroughout the course of the study. *, p<0.05 (Dunnett's).

EXAMPLES Example 1

the MEK inhibitor PD0325901 synergizes with compound A in anantiproliferative assay. The KRAS mutant NSCLC cell line was used in astandard methylene blue proliferation assay. In this assay, compound Aand PD0325901 inhibited the proliferation with a GI50 (that is theconcentration for which 50% of growth inhibition is observed) of 9.41and 36.8 nM respectively (Table 1). When tested in combination, the GI50for BEZ235 decrease to 3.2 nM, that is to say to say that in presence ofPD0325901, 3 times less BEZ235 is required to observe a similar decreasein proliferation. Conversely, the GI50 for PD0325901 decreased to 11.5nM in presence of compound A. The combination Index obtained is of0.718, which correspond to a moderate synergism (Table 2).

TABLE 1 GI50 (in nM) GI50 (in nM) Combination Compound alone incombination Index (CI) Compound A 9.41 4.46 0.718 NVP-LBW624 36.8 11.5

A549 cells were incubated with increasing amount of either compound A orPD0325901 (NVP-LBW624) alone or in combination, for 72 h andproliferation assessed with a standard methylene blue assay. The GI50were determined and Cl quantified with the CalcuSyn software.

TABLE 2 CI Value Phenotypical response 1.1 to 10  antagonism 0.9 to 1.1Nearly additive 0.85 to 0.9  Slight synergism  0.7 to 0.85 Moderatesynergism 0.3 to 0.7 synergim 0.1 to 0.3 Strong synergism

Example 2

the MEK inhibitor and compound A when incubated in cells alone do blocktheir respective downstream effectors (ERK and Akt). When A549 cells areincubated in the presence of both inhibitors, then both pathway areeffectively and simultaneously blocked (FIG. 1).

Example 3

In the BN472 tumor model administration of compound A at a dose of 10mg/kg (p.o., 1 q24 h) or the Raf inhibitor Raf265 (at a dose of 2.5mg/kg, po, &q24 h) produced a weak and not significant antitumoractivity (expressed as T/C) of either 83 and 76. The combination ofcompound A (10 mg/kg) with RAF265, however, produced a strong andstatistically significant antitumor activity with a TIC of 34% (FIG. 2).The combination was well tolerated as no effect on body weight could bemeasured (FIG. 3).

Example 4

KRAS mutant PANC1 cells were grown subcutaneously on the flank of Harlannude mice. Tumor bearing animals were then treated with the indicatedcompounds and dosage regimen in FIG. 4. Evolutions of Tumor volume (A)and body weight (B) were measured throughout the course of the study,until day 12 at which time the animals were sacrificed. *, p<0.05, Oneway ANOVA followed by Tukey post hoc test, showing that the antitumoractivity obtained in the combo group is significantly different from thecontrol and single agent treated groups (synergistic activity betweenthe two compounds).

Example 5

KRAS mutant MiaPaCa2 cells were grown subcutaneously on the flank ofHarlan nude mice. Tumor bearing animals were then treated with theindicated compounds and dosage regimen in FIGS. 5 and 6. Evolutions ofTumor volume (A) and body weight (B) were measured throughout the courseof the study, until day 12 at which time the animals were sacrificed. *,p<0.05, One way ANOVA followed by Tukey post hoc test, showing that theantitumor activity obtained in the combo group is significantlydifferent from the control and single agent treated groups (synergisticactivity between the two compounds).

1. A combination which comprises (a) a phosphoinositide 3-kinaseinhibitor compound inhibitor and (b) a compound which modulates theRas/Raf/Mek pathway, wherein the active ingredients are present in eachcase in free form or in the form of a pharmaceutically acceptable saltor any hydrate thereof, and optionally at least one pharmaceuticallyacceptable carrier; for simultaneous, separate or sequential use.
 2. Acombination according to claim 1, wherein said combination comprises anamount of (a) a phosphoinositide 3-kinase inhibitor compound inhibitorand (b) a compound which modulates the Ras/Raf/Mek pathway in free formor in the form of a pharmaceutically acceptable salt or any hydratethereof in a synergistic ratio.
 3. A combination according to claim 1wherein the phosphoinositide 3-kinase inhibitor compound is COMPOUND A,COMPOUND B or COMPOUND C.
 4. A combination according to claim 1 whereinthe compound which modulates the Ras/Raf/Mek pathway modulates Rafkinase activity.
 5. A combination according to claim 1 wherein thecompound which modulates the Ras/Raf/Mek pathway modulates Mek kinaseactivity.
 6. A combination according to claim 4 wherein the compoundwhich modulates Raf kinase activity is Sorafenib, Raf265, SB590885,XL281 or PLX4032.
 7. A combination according to claim 5 wherein thecompound which modulates Mek kinase activity is PD325901, PD-181461,ARRY142886/AZD6244, ARRY-509, XL518, JTP-74057, AS-701255, AS-701173,AZD8330, ARRY162, ARRY300, RDEA436, E6201, RO4987655/R-7167, GSK1120212or AS703026.
 8. A pharmaceutical composition comprising a quantity whichis jointly therapeutically effective against a proliferative disease ofa combination according to claim 1 and at least one pharmaceuticallyacceptable carrier.
 9. A combination as defined in claim 1 for thetreatment of a proliferative disease.
 10. A combination according toclaim 9 wherein the proliferative disease is a solid tumor.
 11. Acombination according to claim 9 wherein the proliferative disease ismelanoma, lung cancer, colorectal cancer (CRC), breast cancer, kidneycancer such as e.g. renal cell carcinoma (RCC), liver cancer, acutemyelogenous leukemia (AML), myelodysplastic Syndromes (MDS),non-small-cell lung cancer (NSCLC), thyroid cancer, pancreatic cancer,neurofibromatosis or hepatocellular carcinoma.
 12. A combinedpreparation, which comprises (a) one or more unit dosage forms ofphosphoinositide-3 kinase inhibitor and (b) one or more unit dosageforms of a compound which modulates the Ras/Raf/Mek pathway.
 13. Asynergistic combination for human administration comprising a PI3Kinhibitor compound and a compound which modulates the Ras/Raf/Mekpathway, in free form or in the form of a pharmaceutically acceptablesalt or any hydrate thereof, in a w/w combination range whichcorresponds to a synergistic combination range of 10:1 to 1:1 parts byweight in the BN472 tumor model or pancreatic tumor models PANC1 andMiaPaCa2.
 14. A synergistic combination for human administrationcomprising a PI3K inhibitor compound and a compound which modulates theRas/Raf/Mek pathway, in free form or in the form of a pharmaceuticallyacceptable salt or any hydrate thereof, in a w/w combination range whichcorresponds to a synergistic combination range of 1-50 mg/kg and 1-30mg/kg respectively, in the BN472 tumor model or pancreatic tumor modelsPANC1 and MiaPaCa2.
 15. A combination according to claim 1 for humanadministration comprising a synergistic amount of a PI3K inhibitorcompound at about MTD and a compound which modulates the Ras/Raf/Mek at50%-100% of the MTD.